Project Summary/Abstract All organisms obtain the deoxynucleotide substrates for DNA synthesis and repair by the action of an enzyme known as ribonucleotide reductase (RNR). The several known types of RNRs, which have been divided into classes I, II, and III, differ in the transition-metal and free-radical chemistry that they use to initiate their common, challenging reduction/dehydroxylation reaction. Recent studies have shown that many bacteria that infect and cause disease in humans use class I RNRs that differ markedly from the human class I, subclass a enzyme. Some of these microbial RNRs (subclasses b and d) use manganese instead of iron in what is thought to be an adaptation to iron deprivation caused by the human immune response, and others use both metals (subclass c). We just discovered that a new type of RNR from the causative agent of strep throat and scarlet fever may have fully escaped the usual dependence on transition metals by using a previously unknown type of stable amino acid radical, thus founding subclass e. This project will reveal precisely how the members of three new subclasses of class I RNRs (including d and e) that were recently identified in pathogenic bacteria acquire their catalytic activity and initiate nucleotide reduction. The very different initiation chemistry used by the pathogens' enzymes offers opportunities for their selective inhibition by antibiotics. This project will provide the conceptual underpinnings for such drug discovery efforts and will shed light on the ways in which pathogenic microbes have adapted to cope with their hosts' hostile immune response.